Our finding that airway smooth muscle operates over a broad length range suggests 3 related hypotheses: I. Smooth muscle adapts to longer lengths by increasing the number of contractile elements in series. II. Thick-filament lengthening produces the well-known velocity slowing during the rise of activation. III. A thin-filament regulatory mechanism selectively engages filamentous myosin. We have obtained much evidence supporting all three hypotheses, and the results suggest the following Specific Aims to further test these hypotheses and to define the relevant mechanisms: Specific Aim I. Our experiments show that muscles adapt to longer lengths by placing more thick filaments in series, but that the filament increase is only two-thirds of the length increase, suggesting that structures other than thick filaments also contribute to muscle lengthening. Aim I is to quantify the relative contributions of the following four factors likely to increase muscle length: a) longer thick filaments, b) more thick filaments in series, c) lattice tilt, and d) more structures in series with contractile elements. Specific Aim 2. Our experiments show that velocity slowing during the rise of activation is closely correlated with thick-filament formation at an intermediate length and that thick-filament formation is greater at shorter adapted muscle lengths. Aim IIA is to further test the correlation between velocity slowing and filament formation by measuring velocity slowing at different lengths. Aim IIB is to distinguish between 3 possible mechanisms for the slowing, a) longer but fewer thick filaments in series, b) inactivation of whole thick filaments, and c) cytoskeletal rearrangements. Aim IIC will assess the effect of force-altering length manipulations to test whether force changes during the rise of activation alter velocity slowing. Specific Aim 3. The dual role of myosin light-chain phosphorylation, promoting thick-filament formation from myosin monomers and activating myosin[unreadable]s interaction with actin, suggests that another mechanism exists to prevent phosphorylated myosin monomers from interacting with actin before they join filaments. Our observation that calcium promotes crossbridge movement away from thick filaments in the absence of phosphorylation suggests that calcium may regulate this second mechanism. Aim III will use a permeabilized muscle preparation with permanently thiophosphorylated crossbridges to test whether the posited second mechanism exists, how it is regulated, and what other substances affect the interaction of myosin with actin. Relevance. Although asthma and hypertension are usually ascribed to increased smooth muscle tone, the diseases only occur when the muscles are too short. Our finding that smooth muscle undergoes structural adaptations to new lengths suggests that an understanding of the mechanisms of length adaptation would provide a much better understanding of these diseases and suggest new therapeutic targets.